Veterinary World最新文献

Background and aim: Sperm cryopreservation is a cornerstone technology for genetic resource conservation and artificial insemination. However, tropical climatic conditions often compromise post-thaw sperm quality due to enhanced oxidative and thermal stress. This study aimed to optimize glycerol and egg yolk concentrations in a Tris-citrate-glucose (TCG) extender to improve post-thaw sperm quality of Saanen bucks raised in the Mekong Delta, Vietnam.

Materials and methods: The experiment was conducted in two sequential phases. In Phase 1, five glycerol concentrations (4%, 6%, 8%, 10%, and 12%) were tested to identify the optimal permeating cryoprotectant level. In Phase 2, four egg yolk concentrations (5%, 10%, 15%, and 20%) were assessed in combination with the optimal glycerol concentration. Semen from four healthy Saanen bucks was collected using an artificial vagina, diluted in the respective extenders, equilibrated, and frozen in liquid nitrogen. Post-thaw evaluations included overall and progressive motility (phase-contrast microscopy), viability (eosin-nigrosin staining), membrane integrity (hypo-osmotic swelling test), acrosome integrity (Giemsa staining), and DNA fragmentation (sperm chromatin dispersion test). Data were analyzed using one-way analysis of variance with Tukey's post hoc test at p < 0.05.

Results: Glycerol concentration had a significant effect on post-thaw sperm quality (eta squared = 0.93-0.97), with 10% providing the best balance between cryoprotection and cytotoxicity. Incorporating 15% egg yolk with 10% glycerol significantly improved sperm viability (65.5%), membrane integrity (72.3%), and reduced DNA fragmentation (12.4%) compared with other treatments (p < 0.05). Higher glycerol or egg yolk concentrations adversely affected motility due to increased osmotic stress and viscosity.

Conclusion: The combination of 10% glycerol and 15% egg yolk in TCG extender provides optimal cryoprotection for Saanen buck semen under tropical conditions. The resulting post-thaw sperm exhibited high motility, viability, and DNA integrity. This protocol can serve as a region-specific standard for buck semen cryobanking and artificial insemination in tropical climates, supporting genetic improvement and conservation initiatives in Vietnam and other developing regions.

Background and aim: Indigofera zollingeriana is a high-protein tropical legume with potential as a sustainable ruminant feed; however, its ensiling is challenged by rapid proteolysis and ammonia accumulation. Incorporating lactic acid bacteria (LAB) inoculants and natural tannin sources may enhance fermentation quality and nitrogen preservation. This study evaluated the effects of Lactiplantibacillus plantarum inoculant and Acacia mangium leaf tannin extract, individually and in combination, on the chemical composition, ensiling characteristics, in vitro rumen fermentation, digestibility, and metabolomic profiles of Indigofera silage.

Materials and methods: A completely randomized design was used with four treatments: (R0) control, (R1) L. plantarum (10⁶ colony forming units/g FM), (R2) 1% A. mangium extract, and (R3) combination of both additives, each with five replicates. Silages were fermented anaerobically for 30 days. Analyses included proximate composition, pH, lactic acid, ammonia nitrogen (NH₃-N), in vitro gas and methane (CH₄) production, volatile fatty acids (VFA), digestibility, and untargeted metabolomics of both silage and rumen fluid using gas chromatography-mass spectrometry. Data were evaluated using a one-way analysis of variance, Duncan's test, principal component analysis, and partial least squares discriminant analysis.

Results: L. plantarum lowered (p < 0.05) silage pH and fiber fractions but increased NH₃-N due to enhanced deamination. A. mangium tannins effectively suppressed proteolysis, reducing NH₃-N by 11.85%. Their combination improved (p < 0.05) dry- and organic-matter digestibility (↑ ≈ 9%), increased the propionate proportion by 6.82%, and lowered the acetate-to-propionate ratio, indicating a shift toward more energy-efficient rumen fermentation without significant methane (CH₄) inhibition. Metabolomic profiling identified 23 key metabolites in silage and 11 in rumen fluid; acacia tannins increased fatty acyl compounds (+14.3%) while LAB enhanced prenol lipids, reflecting modified lipid and oxygen-derived pathways that improve nutrient utilization.

Conclusion: The combination of L. plantarum and A. mangium leaf extract synergistically improved Indigofera silage quality, nutrient preservation, and digestibility while modulating beneficial metabolites associated with rumen fermentation efficiency. This integrated additive strategy represents a sustainable and locally adaptable approach for tropical ruminant feed production.

Background and aim: Hepatocystis, a malaria-like hemoparasite closely related to Plasmodium, infects non-human primates (NHPs), bats, and other mammals, yet remains understudied in Indonesia. Although Plasmodium detection in primates has been extensively reported, molecular confirmation of Hepatocystis in Indonesian wildlife is lacking. This study aimed to screen NHP fecal samples for Plasmodium infection and to identify any malaria-like parasites using molecular methods.

Materials and methods: A total of 227 fecal samples from captive and rescued NHPs, representing multiple Macaca species and other primates, were collected from Tasikoki Wildlife Rescue Center, Manado, Indonesia, in 2019 and 2021. Genomic DNA was extracted using a QIAamp Fast DNA Stool Mini Kit (Qiagen, Germany) and screened for Plasmodium using polymerase chain reaction (PCR) targeting the mitochondrial small subunit ribosomal RNA gene. Positive amplicons were purified, sequenced, and analyzed using the basic local alignment search tool and phylogenetic reconstruction with MEGA X.

Results: Eight (3.5%) of 227 samples yielded positive PCR bands of approximately 600 bp, differing from the expected 467 bp for Plasmodium. Sequencing of four representative samples (MNig-01, MNig-17, MNig-18, and HM-160) revealed >99.7% identity with Hepatocystis spp. (GenBank: KY653782.1). Multiple sequence alignment confirmed complete nucleotide conservation among the four isolates, and phylogenetic analysis clustered them within the Hepatocystis clade, closely related to Hepatocystis spp. ex Pteropus hypomelanus from Malaysia. All positive detections were from 2019 samples, suggesting temporal variation in infection or vector activity.

Conclusion: This study provides the first molecular evidence of Hepatocystis infection in Indonesian NHPs using fecal DNA, expanding current knowledge of parasite distribution and host range. The successful detection of Hepatocystis through non-invasive sampling underscores the potential of fecal-based PCR for wildlife disease surveillance. These findings highlight the importance of integrating molecular diagnostics into conservation and One Health frameworks to monitor zoonotic parasites and understand host-vector-pathogen interactions in natural ecosystems.

Background and aim: Canine mammary tumors (CMTs) serve as valuable comparative models for human breast cancer (HBC) owing to their shared biological and molecular features. However, well-defined cell lines representing the luminal B subtype remain limited. This study aimed to establish and characterize a novel CMT cell line, designated CMT-622, to expand available in vitro models for luminal B breast cancer research.

Materials and methods: Primary tumor tissue was collected from an 11-year-old female dog diagnosed with high-grade mammary carcinoma (T3N1M0). Tumor cells were isolated using enzymatic digestion and differential adhesion. Morphological, cytogenetic, and immunophenotypic characteristics were assessed using hematoxylin-eosin staining, immunohistochemistry, and immunofluorescence. Growth kinetics, clonogenicity, and chromosomal analyses were performed, and tumorigenicity was evaluated through xenograft assays in nude mice. Drug sensitivity and apoptosis were compared with two existing CMT lines (CMT-1211 and CMT-n7) using the cell counting kit-8 (CCK)-8 assay and flow cytometry.

Results: CMT-622 cells maintained stable proliferation beyond 40 passages with a doubling time of 46.23 h and >15% cloning efficiency. Karyotyping revealed hyperdiploidy (80-110 chromosomes; modal = 87). Immunohistochemistry and immunofluorescence confirmed estrogen receptor (+), progesterone receptor (-), and human epidermal growth factor receptor 2 (weak +) expression, consistent with a luminal B phenotype. Co-expression of cytokeratin-18 and vimentin indicated a partial epithelial-mesenchymal transition (EMT) state. In nude mice, CMT-622 exhibited moderate tumorigenicity and pulmonary metastasis. The line showed intermediate osthole sensitivity (half-maximal inhibitory concentration = 50.48 μM) and an apoptosis rate of 21%, between CMT-1211 and CMT-n7, indicating balanced proliferative and drug-responsive behavior.

Conclusion: CMT-622 represents a newly established luminal B CMT cell line with stable growth, EMT plasticity, and moderate drug sensitivity, reflecting clinically relevant tumor aggressiveness. Its molecular and phenotypic consistency across in vitro and in vivo models underscores its reliability for translational oncology applications. CMT-622 provides a robust preclinical platform for exploring tumorigenesis, metastasis, and therapeutic responses in both veterinary and HBC contexts, bridging comparative and translational cancer research.

Background and aim: Reproductive efficiency during the non-breeding season is a critical factor influencing year-round productivity in sheep farming, particularly in semi-arid environments. Pregnant mare serum gonadotropin (PMSG) is commonly used to induce estrus, yet the optimal dosage that maximizes fertility and lamb growth while minimizing hormonal side effects remains unclear. This study aimed to determine the optimal PMSG dose for enhancing reproductive and growth performance in Dorper × Assaf crossbred ewes under semi-extensive conditions in Palestine.

Materials and methods: A total of 143 non-lactating Dorper × Assaf ewes (aged 2-3 years) were synchronized with intravaginal progestagen sponges for 14 days, followed by intramuscular administration of 400, 500, or 600 international units (IU) of PMSG. Key reproductive traits, estrus response, conception rate, abortion rate, fecundity, and lamb survival, were recorded, alongside lamb birth and weaning weights. Data were analyzed using a one-way analysis of variance and a general linear model incorporating parity, litter size, and sex effects, with significance set at p < 0.05.

Results: High conception and lambing rates were observed across all groups. The 400 IU PMSG dose resulted in the lowest abortion rate (2.44%) and highest lamb survival (96.9%), while higher doses (500-600 IU) increased abortion incidence without improving fertility outcomes. Birth weight increased with PMSG level (3.42-3.83 kg), whereas weaning weight peaked at 500 IU (22.18 kg). Litter size and lamb sex significantly affected both birth and weaning weights, with singletons and males being heavier.

Conclusion: Administering 400 IU of PMSG provides the optimal balance between reproductive performance, lamb survival, cost-efficiency, and animal welfare. Excessive hormonal stimulation at higher doses offers no additional reproductive benefit and may compromise ewe health. Implementing this moderate, breed-specific hormonal protocol enhances fertility while reducing veterinary intervention and production costs, promoting sustainability and welfare-conscious management. These outcomes directly contribute to sustainable development goals (SDG) 2 - zero hunger and SDG 12 - responsible consumption and production, advancing resilient and ethical small-ruminant farming systems.

Background and aim: Milk yield variability in tropical dairy goats is driven not only by nutrition but also by complex metabolic and hormonal regulation. Conventional nutrition studies often overlook the physiological mechanisms underlying lactation efficiency. This study aimed to integrate metabolomic and hormonal analyses to identify biomarkers associated with high and low milk yield performance in Sapera goats. It provides the first untargeted ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS)-based metabolomics characterization linking metabolic and endocrine profiles to lactational efficiency in tropical dairy systems.

Materials and methods: Twenty lactating Sapera goats were categorized into two groups: High-yielding (HY) (>1000 mL/day) and low-yielding (LY) (≤1000 mL/day). All animals were fed identical diets formulated according to National Research Council (2007) standards and managed under uniform housing conditions. Milk composition, blood metabolites (glucose, cholesterol, total protein, and triglycerides), and plasma hormones (leptin and insulin-like growth factor-1) were quantified using colorimetric and enzyme-linked immunosorbent assay assays. Untargeted metabolomics of milk samples was performed using UHPLC-HRMS. Metabolites were identified through multi-database annotation, and statistical comparisons were conducted using independent t-tests with p < 0.05.

Results: Milk composition did not differ significantly between groups (p > 0.05). However, HY goats exhibited higher glucose and cholesterol concentrations and lower leptin levels (2.39 ± 0.42 ng/mL vs. 3.00 ± 0.44 ng/mL). Metabolomic analysis identified 213 metabolites, 19 unique to HYs, 28 to LYs, and 166 metabolites were found in both groups. HY goats showed enrichment of metabolites linked to lipid metabolism, membrane synthesis, and antioxidant defense (e.g., uric acid and phosphoserine derivatives), while LY goats displayed compounds indicative of metabolic stress and detoxification load (e.g., glycocholic acid and 3-furoic acid). Integrative correlation mapping revealed coordinated regulation between blood and milk metabolites in HY animals.

Conclusion: HY and LY goats possess distinct metabolic and hormonal signatures despite similar milk composition. Identified biomarkers such as uric acid and glycerophosphoserine highlight pathways supporting efficient nutrient utilization and milk synthesis. These findings provide a foundation for precision feeding and biomarker-guided selection strategies to enhance productivity and sustainability in tropical dairy goat systems.

Background and aim: The giant freshwater prawn (Macrobrachium rosenbergii) is an economically valuable aquaculture species, yet its production often faces challenges related to poor growth and disease outbreaks caused by intensive farming practices and excessive antibiotic use. Probiotics offer a sustainable alternative for improving growth, immunity, and pond health, but their efficacy and safety are highly strain specific. This study aimed to evaluate the probiotic potential of Bacillus pseudomycoides, Bacillus safensis, and Bacillus infantis isolated from prawn farms in Kalasin Province, Thailand, focusing on digestive enzyme activities, hemolytic safety, and antibiotic susceptibility.

Materials and methods: Three Bacillus strains previously identified by 16S ribosomal RNA sequencing were characterized using standard in vitro assays. Amylase, protease, and lipase activities were assessed using the halo/colony (H/C) ratio and enzyme unit measurements. Hemolytic patterns were examined on Columbia agar supplemented with 5% sheep blood, and antibiotic susceptibility was tested by the disk diffusion method against seven antibiotics following Clinical and Laboratory Standards Institute (2024) guidelines. All assays were performed in triplicate, and data were analyzed using one-way analysis of variance (p < 0.05).

Results: All strains exhibited strong amylolytic (H/C ratio 2.05-2.27) and proteolytic (H/C ratio 1.75-1.96) activities, while lipase activity was undetectable. Hemolysis testing revealed γ-hemolysis for all strains, confirming non-hemolytic and non-pathogenic properties. Antibiotic susceptibility profiles indicated broad sensitivity to penicillin G, tetracycline, streptomycin, erythromycin, and chloramphenicol. Moderate susceptibility to vancomycin was observed in B. pseudomycoides and B. safensis, while B. infantis remained fully susceptible. The results suggest a strong digestive enzyme potential and an acceptable safety profile among all isolates.

Conclusion: The evaluated Bacillus strains exhibit favorable probiotic attributes, including high amylase and protease activity, non-hemolytic safety, and broad antibiotic susceptibility, which support their suitability for probiotic application in M. rosenbergii culture. These native isolates may serve as sustainable, locally adapted alternatives to antibiotics, contributing to improved feed efficiency, growth, and disease resistance in freshwater prawn aquaculture. Further in vivo validation and genomic safety analyses are recommended to confirm their efficacy under commercial conditions.

Background and aim: Growth retardation syndrome in cultured Penaeus vannamei has been associated with Enterocytozoon hepatopenaei (EHP) and a recently identified decapod hepanhamaparvovirus (DHPV) genotype V. However, data on its prevalence, pathogenicity, and interaction with the shrimp hepatopancreatic microbiome in Thailand remain limited. This study aimed to determine the incidence and co-infection rate of DHPV genotype V with EHP, evaluate its pathogenic potential, and explore microbiome alterations associated with infection.

Materials and methods: Between 2022 and 2023, 1,270 shrimp from 127 grow-out ponds across 46 farms in eastern Thailand and post-larvae 12 from five hatcheries in the south were screened for DHPV and EHP by polymerase chain reaction. Six representative isolates underwent phylogenetic analysis based on non-structural protein 1 (NS1) and NS2 genes. Pathogenicity was evaluated by immersion challenge bioassays in specific pathogen-free P. vannamei. Hepatopancreatic microbiomes of naturally infected and healthy shrimp were compared using 16S ribosomal RNA gene sequencing and Quantitative Insights Into Microbial Ecology 2-based analysis.

Results: DHPV was detected in 54.33% (69/127) of ponds and 4% (1/25) of hatchery tanks. Co-infection with EHP occurred in 40.16% of ponds. Phylogenetic analysis showed 97.99%-98.82% similarity with DHPV genotype V from South Korea, confirming transboundary genetic relatedness. Experimental infection caused low mortality (20%) but resulted in viral replication (10¹-10³ copies/μL) and characteristic intranuclear inclusion bodies in hepatopancreatic cells. DHPV-infected shrimp exhibited distinct microbiome profiles with elevated Firmicutes, Planctomycetota, and Actinobacteriota abundances, supporting a pathobiome shift during infection.

Conclusion: This is the first report of DHPV genotype V in P. vannamei from Thailand and its frequent co-infection with EHP. Despite its low experimental virulence, the widespread occurrence and microbiome dysbiosis suggest that it may have subclinical impacts that could exacerbate growth retardation. Routine molecular screening in hatcheries and farms, coupled with integrated viral-microbiome surveillance, is essential for sustainable aquaculture biosecurity and aligns with the United Nations Sustainable Development Goal 14 (Life Below Water) by promoting resilient aquatic food systems.

Background and aim: Conventional egg-yolk and milk-based extenders are widely used for semen cryopreservation but pose biosafety concerns and compositional variability that compromise standardization. Liposome technology offers a biosecure, uniform alternative. This study aimed to develop soy lecithin-derived nanoliposomes (NLs) using an ultrasonic-based process and to evaluate their efficacy as a Tris-based extender for Ongole-grade bull semen cryopreservation.

Materials and methods: Soy lecithin NLs were prepared through probe ultrasonication (15-45 min) and ultracentrifugation, followed by physicochemical characterization using particle size analysis, scanning electron microscopy (SEM), and high-resolution transmission electron microscopy. Tris-NL (TNL) extenders were formulated at concentrations of 5-25 mg/mL and compared with a Tris-egg-yolk (TEY, 20%) control. Fresh semen from five Ongole-grade bulls was evaluated for motility, viability, and morphology before and after freezing. Kinematic parameters were assessed through computer-assisted semen analysis, plasma membrane integrity by hypoosmotic swelling test, acrosome integrity using fluorescein isothiocyanate-conjugated peanut agglutinin/propidium iodide staining, and DNA fragmentation by Halomax-sperm chromatin dispersion assay.

Results: Optimized sonication (45 min) produced stable NLs (mean diameter 76 nm, zeta potential -43.2 mV) with uniform spherical morphology. Among the tested formulations, TNL 5-10 mg/mL showed significantly higher (p < 0.05) post-equilibration motility (up to 98%), progressive motility, and kinematic parameters (velocity curved line, velocity average path, and velocity straight line) than TEY. Post-thaw evaluations demonstrated improved sperm viability (≈66%), reduced abnormalities (<7%), enhanced plasma-membrane and acrosomal integrity, and lower DNA fragmentation (~1.2%) in the 5-10 mg/mL groups. SEM confirmed smoother sperm surfaces with minimal cryo-damage compared with TEY.

Conclusion: Soy lecithin-derived NLs at 5-10 mg/mL serve as an effective and biosecure substitute for egg yolk in Tris extenders, enhancing motility, viability, and structural integrity of Ongole-grade bull spermatozoa. This locally developed, plant-based nanotechnology supports biosafety, import substitution, and sustainability of artificial insemination programs in Indonesia.

Background and aim: Cryopreservation is an essential assisted reproductive technology for preserving valuable animal genetics; however, it induces osmotic and oxidative stress that compromises sperm quality. Melatonin (MLT) functions as both an antioxidant and a signaling molecule through specific membrane receptors, melatonin 1 (MT1) and melatonin 2 (MT2). This study aimed to determine the association between sperm freezability, MLT receptor expression on spermatozoa, and MLT concentration in the seminal plasma of goat bucks.

Materials and methods: Semen samples were collected weekly from seven clinically healthy Saanen bucks (2-3 years) maintained under natural photoperiod and standardized feeding conditions. A total of 124 ejaculates were analyzed seasonally (spring-winter). Sperm quality parameters, including motility, viability, capacitation status (chlortetracycline [CTC] assay), plasma membrane fluidity, and acrosome integrity, were evaluated before and after cryopreservation. Expression levels of MT1 and MT2 receptors were quantified by Western blot, while MLT concentration in seminal plasma was measured by enzyme-linked immunosorbent assay. Pearson's correlation and determination coefficients (R, R²) were computed to assess associations between sperm quality, receptor expression, and MLT concentration.

Results: Western blot analysis revealed variable MT1 (16 kDa) and MT2 (28-75 kDa) expression throughout the year. High negative correlations (R > -0.9, R² > 0.8) were observed between the expression of both receptors and the proportion of acrosome-reacted (AR) spermatozoa (CTC-AR pattern), except for MT2 (75 kDa). Conversely, seminal plasma MLT concentration showed a strong positive correlation (R > 0.9, R² > 0.8) with capacitated sperm having intact acrosomes (CTC-B pattern). Seasonal variation in receptor expression, rather than MLT concentration, influenced sperm cryoresistance.

Conclusion: Sperm freezability in goats is closely linked to the expression of MLT receptors and seminal plasma MLT concentration. Higher receptor expression corresponds to improved post-thaw sperm quality, likely by mitigating cryocapacitation and acrosomal damage. These findings suggest that enhancing MLT receptor expression or modulating photoperiodic exposure could optimize semen cryopreservation protocols and reproductive efficiency in caprine species.